Diagnosis of enzootic pneumonia in Danish cattle: reverse transcription-polymerase chain reaction assay for detection of bovine respiratory syncytialvirus in naturally and experimentally infected cattle

A reverse transcription-polymerase chain reaction (RT-PCR) assay was develo ped for detection of bovine respiratory syncytial virus (BRSV) in lung tiss ue of naturally and experimentally infected cattle. Primers were selected f rom the gene coding the F fusion protein, which is relatively conserved amo ng BRSV isolates. The RT-PCR assay was highly specific, it yielded positive reactions only when performed on BRSV-infected cell cultures or tissues. T he detection limit of the RT-PCR assay was assessed as 5 TCID50. BRSV was d etected in tissues of the respiratory tract and in the tracheobroncheal lym ph node of calves euthanized 2-8 days after experimental infection with BRS V, whereas samples of other tissues and samples from mock-infected animals were negative at all time points. Examination of lung samples from 8 differ ent regions of the lungs revealed that although the virus was most often fo und in the cranioventral lobules, it was frequently present in all lung lob ules. Microbiologic examinations of all acute fatal cases of pneumonia (135 animals) in cattle submitted for diagnostic purposes during 1 year reveale d that Actinomyces pyogenes (11%), Haemophilus somnus (10%). Pasteurella sp . (7%), and Pasteurella haemolytica (7%) were the most common bacterial age nts found in the lungs. BRSV was identified using a conventional antigen en zyme-linked immunosorbent assay (ELISA) in 23 (17%) animals. The establishe d BRSV-specific RT-PCR assay yielded positive results for the same 23 anima ls, in addition, 10 animals that were negative with the ELISA were positive with the RT-PCR assay. These results indicates that the RT-PCR assay can b e a sensitive, reliable alternative to conventional diagnostic procedures.